Gustavo Mostoslavsky, M.D., Ph.D.
Department of Medicine
Embryonic Stem Cell Modeling of Intestinal Differentiation
Embryonic Stem Cells (ESC) are pluripotent undifferentiated cells capable of giving rise to cells from all three germ layers. This unique ability makes them ideal candidates to model early development allowing us to study the basic signaling mechanisms involved in stem cell fate determination. At the same time, manipulating ESC differentiation toward a specific developmental pathway holds a great promise for their use in regenerative medicine. One focus of our lab is differentiating mouse ESC into intestinal epithelial cells in order to understand the complex signaling pathways involved in intestinal commitment from endodermal progenitors and undifferentiated stem cells.
Our lab has a major interest in the study of induced Pluripotent Stem cells or iPS cells and the development of tools for their generation and characterization. Recent pioneering work by the laboratory of Dr. Yamanaka showed that fibroblasts transduced with retroviral vectors expressing four transcription factors, Oct4, Klf4, Sox2 and cMyc can be reprogrammed to become pluripotent stem cells that appear almost indistinguishable from ESC. In contrast to ESC, iPS cells are genetically identical to the individual from whom they are derived, raising the prospect of utilizing iPS cells for autologous cell based therapies without risk of rejection. We have recently developed a single lentiviral vector, named pHAGE-STEMCCA, capable of generating iPS cells from post-natal fibroblasts with the highest efficiency reported to date. We aimed at using iPS cells in parallel to ESC for the study of intestinal lineage specification and their potential for regenerative medicine.
Characterization and Isolation of Intestinal Stem Cells
The identification of Intestinal Stem Cells (ISCs) has long-eluded investigators. The recent discovery of LGR5 as a putative marker of ISCs has opened a window for their study and characterization. We use several methods, including gene marking and gene transfer technologies to study ISC biology and their potential use in cell and gene therapy.
Hematopoietic Stem Cell Manipulation for the Study of Stem Cell Self-Renewal and Differentiation
Hematopoietic Stem Cells (HSCs) are the most thoroughly characterized stem cell population in the body and their study has resulted in well established methods for their isolation, purification and reliable assays of HSC function. During the last few years we have substantially improved our ability to genetically manipulate HSCs using viral vectors for gene transfer. Despite these efforts, few genes are known to play a role in the processes of stem cell self-renewal and differentiation. Understanding the molecular mechanisms that govern those unique functions are crucial for developing the promise that stem cells hold for developmental biology and regenerative medicine. In our lab, we use lentiviral viral gene transfer to study the role of several molecules in long-term HSC self-renewal and differentiation.
Hematopoietic Stem Cell Manipulation for the Correction of Immunodeficiencies
Our longstanding interest in the immune system combined with our experience in manipulating HSCs have culminated in several studies whose goal is genetic correction of Severe Combined Immunodeficiency (SCID). It has recently become clear that many SCID patients suffer from a spectrum of previously unrecognized hypomorphic mutations leading to partially impaired V(D)J rearrangement activity. The best example of this type of immunodeficiency is Omenn Syndrome (OS), which is caused in most cases by Rag hypomorphic mutations. While it is well established that the genetic defect in either of the RAG genes is the first determinant of the clinical presentation, the mechanism by which specific Rag mutations induce such diverse immunological phenotypic outcomes is still poorly understood. We have recently started a collaboration with a group at Harvard Medical School to use a variety of lentiviral vectors expressing Rag1 to study its role in immune dysregulation and to develop a new therapeutic approach for Rag1 related immunodeficiency based on lentiviral mediated gene therapy.
1. Mostoslavsky, G., Kotton, D.N., Fabian, A.J., Lee, J-S, Gray, J.T., and Mulligan, R.C. Efficiency of transduction of highly purified murine hematopoietic stem cells by lentiviral and oncoretroviral vectors under conditions of minimal in vitro manipulation. 2005. Mol. Therapy. 11 (6): 932-940.
2. Mostoslavsky, R., Chua, K.F, Lombard, D.L., Pang, W.W, Fischer, M.R., Gellon, L., Liu, P., Mostoslavsky, G., Franco, S., Murphy, M.M., Mills, K.D., Patel, P., Hsu, J., Hong, A.L., Ford, E., Cheng, H-L., Kennedy, C., Nunez, N., Bronson, R., Frendewey, D., Auerbach, W., Valenzuela, D., Karow, M., Hottiger, M.O., Hursting, S., Barrett, J.C., Guarente, L., Mulligan, R., Demple, B., Yancopolous, G.D., and Alt, F.W. Genomic instability and aging-like phenotype in the absence of mammalian SIRT6. 2006. Cell. 124:1-15.
3. Murphy, G.J., Mostoslavsky, G., Kotton, D.N. and Mulligan R.C. Exogenous control of mammalian gene expression via modulation of translational termination. 2006. Nat. Medicine. 12(9): 1093-1099. doi:10.1038/nm1376.
4. Mostoslavsky G., Fabian, A.J., Rooney, S., Alt, F.W. and Mulligan, R.C. Complete correction of murine Artemis immunodeficiency by lentiviral vector-mediated gene transfer. 2006. PNAS. 103(44): 16406-16411. Doi:10.1073/pnas.0608130103.
5. De Coppi, P., Bartsch, G. Jr., Siddiqui M.M., Xu, T., Santos, C.C., Perin, L., Mostoslavsky, G., Serre, A.C., Snyder, E.Y., Yoo, J.J., Furth, M.E., Soker, S. and Atala, A. Isolation of amniotic stem cell lines with potential for therapy. 2007. Nat. Biotech. 25(1): 100-106. Doi:10.1038/nbt1274.
6. Pan, H., Mostoslavsky, G., Eruslanov, E., Kotton, D.N., and Kramnik, I. Dual-promoter lentiviral system allows inducible expression of noxious proteins in macrophages. J. Immunol. Methods. 2008. Jan 1;329(1-2):31-44. Epub 2007 Oct 18.
7. Sommer, C., Stadtfeld, M., Murphy, G.J., Hochedlinger, K., Kotton, D.N., and Mostoslavsky, G. iPS Cell Generation Using a Single Lentiviral Stem Cell Cassette. 2008. Stem Cells. Epub Dec 18 2008.